ClinVar Genomic variation as it relates to human health

This sequence change replaces alanine, which is neutral and non-polar, with threonine, which is neutral and polar, at codon 364 of the MYBPC3 protein (p.Ala364Thr). This variant also falls at the last nucleotide of exon 12, which is part of the consensus splice site for this exon. This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individuals with hypertrophic cardiomyopathy (PMID: 20513729, 21239446, 21302287, 25740977). ClinVar contains an entry for this variant (Variation ID: 454301). An algorithm developed to predict the effect of missense changes on protein structure and function (PolyPhen-2) suggests that this variant is likely to be tolerated. Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). Algorithms developed to predict the effect of sequence changes on RNA splicing suggest that this variant may disrupt the consensus splice site. For these reasons, this variant has been classified as Pathogenic.

The c.1090G>A variant (also known as p.A364T), located in coding exon 12 of the MYBPC3 gene, results from a G to A substitution at nucleotide position 1090. The threonine at codon 364 is replaced by alanine, an amino acid with similar properties. However, this change occurs in the last base pair of coding exon 12 and may have some effect on normal mRNA splicing. This alteration has been reported in several individuals with hypertrophic cardiomyopathy (HCM) (Melacini P et al. Eur. Heart J., 2010 Sep;31:2111-23; Millat G et al. Clin. Chim. Acta, 2010 Dec;411:1983-91; Fokstuen S et al. J. Med. Genet., 2011 Aug;48:572-6; Roncarati R et al. J. Cell. Physiol., 2011 Nov;226:2894-900; Calore C et al. J. Med. Genet., 2015 May;52:338-47). RNA and minigene studies indicate that this alteration results in abnormal splicing (Crehalet H et al. Cardiogenetics, 2012; v.2:e6; Singer ES et al. Circ Genom Precis Med. 2019 Jan;12(1):e002368). Both the nucleotide and amino acid positions are well conserved in available vertebrate species. In silico splice site analysis predicts that this alteration may weaken the native splice donor site. In addition, as a missense substitution, this alteration is predicted to be tolerated by in silico analysis. This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). Based on the majority of available evidence to date, this variant is likely to be pathogenic.

The MYBPC3 c.1090G>A variant is predicted to result in the amino acid substitution p.Ala364Thr. This variant is located at the last nucleotide of exon 12 and is predicted to alter splicing based on available splicing prediction programs (Alamut Visual Plus v1.6.1). This variant was reported in numerous individuals with hypertrophic cardiomyopathy or dilated cardiomyopathy (Melacini et al. 2010. PubMed ID: 20513729; Millat et al. 2010. PubMed ID: 20800588; Roncarati et al. 2011. PubMed ID: 21302287; Calore et al. 2015. PubMed ID: 25740977; Table S3, Mazzarotto et al. 2020. PubMed ID: 31983221; Table S1, Preveden et al. 2022. PubMed ID: 35208637). mRNA studies using affected individual's blood sample showed that this variant results in the skipping of exon 12 and a subsequent frameshift (Figure S1F, Singer et al. 2019. PubMed ID: 30645170). This variant has not been reported in a large population database (http://gnomad.broadinstitute.org), indicating this variant is rare. This variant is interpreted as pathogenic.

This variant changes the last nucleotide c.G of exon 12 of the MYBPC3 gene and is predicted to impair RNA splicing at the intron 12 splice donor site. RNA studies from a carrier individual have shown that this variant causes an out-of-frame skipping of exon 12 (PMID: 30645170, 33530161). This variant is expected to result in an absent or non-functional protein product. This variant has been reported in at least 7 individuals affected with hypertrophic cardiomyopathy (PMID: 20513729, 20800588, 21239446, 21302287, 25740977, 30645170, 32481709, 32841044, 35208637; SCV000924849.1), and in one individual affected with dilated cardiomyopathy (PMID: 31983221). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Loss of MYBPC3 function is a known mechanism of disease (clinicalgenome.org). Based on the available evidence, this variant is classified as Pathogenic.

This variant changes the last nucleotide c.G of exon 12 of the MYBPC3 gene and is predicted to impair RNA splicing at the intron 12 splice donor site. RNA studies from a carrier individual have shown that this variant causes an out-of-frame skipping of exon 12 (PMID: 30645170, 33530161). This variant is expected to result in an absent or non-functional protein product. This variant has been reported in at least 7 individuals affected with hypertrophic cardiomyopathy (PMID: 20513729, 20800588, 21239446, 21302287, 25740977, 30645170, 32481709, 32841044, 35208637; SCV000924849.1), and in one individual affected with dilated cardiomyopathy (PMID: 31983221). This variant has not been identified in the general population by the Genome Aggregation Database (gnomAD). Loss of MYBPC3 function is a known mechanism of disease (clinicalgenome.org). Based on the available evidence, this variant is classified as Pathogenic.

Testing for our patient was performed at Invitae. Given the moderate case data, possible effect on splicing, and absence in general population databases, we consider this variant likely pathogenic and we do feel it is suitable for assessing risk in healthy relatives ("predictive genetic testing"). The variant has been seen in at least 7 unrelated cases of HCM, one of which is a patient at our center. There is moderate case data. All of the cases are of Italian descent. Roncarati et al (2011) saw this variant in 1 patient with HCM and noted there may be a potential effect on splicing. Patient was recruited in Italy, but not in Padua. Calore et al. (2015) reports this variant in 5 index cases of HCM from a cohort in Padua, Italy. The variant has also been reported in other papers from overlapping groups in Italy (Fokstuen 2011; Melacini 2010), but these cases are not counted in the overall case count given possible redundancy of patients. The paper by Roncarati et al. also cites Smaniotto et al. (2009), but I was unable to find the actual paper. Even so, the authors were Italian and any cases reported in that paper could be overlapping. Per the lab report, algorithms predict that this variant could have an effect on splicing. Splicing and other protein-truncating variants in MYBPC3 are a frequent cause of HCM (Erdmann et al. 2001 & 2003; Stenson et al 2003; Harvard Sarcomere Protein Gene Mutation Database). Many MYBPC3 splice variants have been reported in association with HCM including IVS2-1G>A, IVS6-2A>C IVS7+1G>A, IVS8+1G>A, IVS12-2A>G, IVS14-2A>G, IVS16-1G>A, IVS22+1G>A, IVS24+1G>A, IVS28+1G>A, IVS32+1G>A, and IVS33+1G>A (Harvard Sarcomere Protein Gene Mutation Database). Furthermore, these types of variants in MYBPC3 are not seen in individuals without cardiomyopathy (Pan et al 2012). Another variant at the same codon Ala364Pro, which is considered a VUS by a reputable laboratory. That variant is absent from gnomAD. The variant is not listed in the Genome Aggregation Consortium Dataset (gnomAD; http://gnomad.broadinstitute.org/), which currently includes variant calls on >140,000 unrelated individuals of African, Asian, European, Latino, and Ashkenazi descent. The average coverage at that site in gnomAD genomes is 51x and in gnomAD exomes is 32x.